GLUTATHIONE TRANSFERASES AND HERBICIDE DETOXIFICATION IN SUSPENSION-CULTURED CELLS OF GIANT FOXTAIL (SETARIA-FABERI)

Glutathione transferases (GSTs) catalysing the conjugation of 1-chloro -2,4-dinitrobenzene, the chloro-s-triazine herbicide atrazine, the chl oroacetanilide herbicides metolachlor and alachlor and the diphenyl et her herbicide fluorodifen have been identified in suspension-cultured cells derived from the grass weed giant foxtail (Setaria faberi Herrm. ). In contrast to suspension-cultured cells of maize, where atrazine-c onjugating GSTs are lost during de-differentiation, the GSTs active to ward this herbicide in S. faberi plants were also expressed in culture s, suggesting that these isoenzymes are subject to different regulatio n in the crop and weed. As a result, glutathione conjugation was the m ajor route of atrazine metabolism in S. faberi cultures. Activities of these GSTs were maximal three days after sub-culturing when the cells were dividing most actively, when they were determined to be in the o rder CDNB > alachlor > metolachlor = fluorodifen > atrazine. This indi cated that GSTs which are enhanced during cell division can metabolise herbicides. On the basis of activity per mg protein, GST activities i n the cultures were between 20 and 60-fold higher than those determine d in the foliage of S. faberi seedlings. The GSTs with activity toward s CDNB were resolved into three peaks following anion-exchange chromat ography at pH 7.8 using Q-Sepharose. Peak 1 GSTs were not retained, wh ile peak 2 and peak 3 were sequentially resolved with an increasing co ncentration of salt. Peak I GSTs showed activity toward metolachlor an d atrazine but showed little activity toward fluorodifen. Peak 2 and p eak 3 GSTs were active toward atrazine and metolachlor, with peak 3 be ing particularly associated with activity toward fluorodifen. The GSTs in these peaks were then further purified using S-hexyl-glutathione-a garose affinity chromatography. In each case, the affinity-bound fract ion of the GSTs consisted of 28 kDa and 26 kDa polypeptides, suggestin g that the GST isoenzymes in S. faberi cultures are composed of relate d subunits. Our results demonstrate that the GST isoenzymes involved i n herbicide metabolism in suspension cultures of a grass weed show a s imilar level of complexity to that determined in maize cell cultures. (C) 1998 SCI.